DESCRIPTION (Applicant's abstract): Cardiovascular disease is responsible for over 50 percent of the deaths in the United States. Plaque formation, a hallmark of atherosclerosis, predictably occurs at arterial sites characterized by shear stress that is low, high or alternating. The overall objective of this research is to determine the molecular mechanisms by which shear stress participates in endothelial injury and, thus, contributes to the pathogenesis of atherosclerosis. As mediators of cell-cell cohesion and communication, endothelial adherens junctions represent a potential vulnerability to shear stress perturbations. This investigation centers on the junctional role(s) of intra-and intercellular signaling, focusing on the Wnt pathway. The theory that adherens junctions and Wnt signaling are linked stems from the functional roles of beta-catenin as a pivotal protein in cell cohesion and as a key regulatory molecule in the Wnt pathway. Preliminary studies have demonstrated that in subconfluent endothelial cells, about-catenin is translocated to the nucleus, an event associated with Wnt pathway activation. In addition, pulsatile but not steady shear stress results in a marked decrease in Wnt-1 cell labeling. The specific aims are: (1) to determine the phosphorylation state and a subcellular compartmentalization of beta-catenin in endothelial cells for compatibility with an operative Wnt signaling pathway, (2) to test the hypothesis that shear stress modulates Wnt signaling in endothelial cells utilizing a TCF/LEF reporter gene construct and (3) to identify the mechanisms by which shear stress may modulate Wnt signaling in endogenous endothelium. Methods to accomplish these aims will include the use of an in vitro shear stress apparatus, Western blotting, RT-PCR, reporter gene constructs, cell transfection, high resolution 3-D microscopy and immunocytochemistry. These proposed studies would establish a linkage between shear stress as a controlling influence on adherens junctions in endothelium and that a consequence of this regulation is modulation of Wnt siqnaling.